2010-2014 has been a productive period. Our success in devising novel methods to study the molecular mechanisms underlying cardioprotection has exceeded expectations. We achieved four major accomplishments, all of which were driven by technological innovations. Briefly, we created novel platforms to analyze quantitative protein phosphorylation and protein turnover rates; we expanded the reach of 'omics techniques to a widening field of novel proteome parameters; we built novel computational tools and data-to- knowledge workflows (Cardiac Organellar Protein Atlas Knowledgebase (COPaKB) and ProTurn) and integrated them into our experimental designs and thought processes. These endeavors are highly significant because (1) through them we gained an elevated understanding into the many aspects of cardiac mitochondrial functions within the purview of cardioprotection; and (ii) these new tools pave the way for future investigations and the testing of new hypotheses in multifarious directions. The major scientific outputs supported by this Award are highlighted by a series of 11 peer-reviewed publications in journals such as Circ Res, J Clin Invest, and J Proteome Res (the Ping laboratory has contributed 36 publications from 2011- 2014). Furthermore, the MERIT Award supported 24 trainees. The trainees collectively received 11 academic honors and recognitions. The PI also received 3 international awards, participated in 6 NIH workshops, co- authored 4 white papers/ reviews, and succeeded in 1 patent application (UC 2013-137-0). In the extension period, we will examine the spatiotemporal dynamics of mitochondrial cardioprotective signaling pathways. Innovation will maintain priority in the MERIT Award, as we will continue to create new in- house technologies and apply them directly to elucidate the regulatory mechanisms of cardiac disease and protection. Three Specific Aims are proposed in this renewal. First, we will elucidate the spatial distribution of mitochondrial protein complex assembly and interaction in the setting of cardioprotection and/or elevated oxidative stress. Second, we will examine the effect of cardioprotection and/or oxidative stress on protein temporal dynamics. Third, we will design and construct an 'omics-based screening strategy to select human induced-pluripotent stem cell (iPSC)-derived cardiomyocytes that are resistant to oxidative stress and injury. We will validate molecular markers and evaluate their informativeness in predicting stress-resistant phenotypes in subsequent lines. We are convinced that these studies will propel our knowledge on cardioprotective environments and also avail the use of iPSC-derived cardiomyocyte experimental models in future discoveries and therapies. RELEVANCE (See instructions): Myocardial ischemic injury affects millions of people worldwide and is a leading cause of death. The proposed studies will examine the mitochondrial signaling mechanisms of NO-donor or PKCe-mediated cardioprotection against myocardial ischemic injury. Understanding the pathogenic mechanisms of and protection from ischemic injury has major implications to improve human health, and will create numerous opportunities to make key contributions to the fundamental knowledge of mitochondria and survival signaling using new technologies.